Lab part 2

Structures and Functions of Microtubules

Microtubules are filamentous intracellular structures
that are responsible for various kinds of movements
in all eukaryotic cells. Microtubules are involved
in nucleic and cell division, organization of
intracellular structure, and intracellular transport,
as well as ciliary and flagellar motility. Because
the functions of microtubules are so critical
to the existence of eukaryotic cells (including
our own), it is important that we understand
their composition, how they are assembled and
disassembled, and how their assembly/disassembly
and functions are regulated by cells.

For the sake of brevity, only the very basic
and universal concepts about microtubules and
their organization into flagella will be presented
here, leaving many questions unanswered. You
will find that textbooks provide more complete
descriptions of microtubules and their structures
and functions, but they also leave many questions
unanswered. Textbooks seldom tell us is how much
science knows and does not know about them, and
of course they cannot be up to date with the
latest discoveries. To fully understand a subject
it is important to go to multiple sources. If
the subject is especially important to you, you
should seek the primary literature, namely original
research reports.

"Building blocks" of microtubules
- tubulins

All eukaryotic cells produce the protein tubulin,
in the usual way. The usual way, of course, is
by transcription of genes coding for tubulin
to produce messenger RNA, followed by the translation
of mRNA by the ribosomes in order to produce
protein. Cells maintain at least two types of
tubulin, which we call alpha tubulin and beta
tubulin. However, it is doubtful that the two
types can found in cells as individual proteins.

Alpha and beta tubulin spontaneously bind one
another to form a functional subunit that we
call a heterodimer. A heterodimer is a
protein that consists of twodifferent
gene products. The term is entirely descriptive
- the prefix hetero- means "different," the
prefix di- means "two," and
the suffix -mer refers to a unit, in this
case a single polypeptide. Obviously, cells do
not continue to make tubulin (or any other protein)
until they run out of resources. Some process
must regulate the synthesis of tubulin. A common
regulatory mechanism is feedback inhibition.

The figure illustrates the inhibition
of tubulin synthesis by the presence of heterodimers
in the system. Exactly how that inhibition takes
place is irrelevant to this discussion. More
about the important concept of feedback inhibition
can be found elsewhere.

Assembly of microtubules

When intracellular conditions favor assembly,
tubulin heterodimers assemble into linear protofilaments.
Protofilaments in turn assemble into microtubules.
All such assembly is subject to regulation by
the cell.

Microtubules form a framework for structures
such as the spindle apparatus that appears during
cell division, or the whiplike organelles known
as cilia and flagella. Cilia and flagella are
the most well-studied models for microtubule
structure and assembly, and are often used by
textbooks to introduce microtubules.

Dynamic instability of microtubules

Under steady state conditions a microtubule
may appear to be completely stable, however there
is action taking place constantly. Populations
of microtubules usually consist of some that
are shrinking and some that are growing. A single
microtubule can oscillate between growth and
shortening phases. During growth, heterodimers
are added on to the end of a microtubule, and
during shrinkage they come off as intact subunits.
The same heterodimer can come off and go back
on.

Since even apparently stable microtubular
structures have an intrinsic instability, they
are considered to be in a dynamic equilibrium,
or steady state. Look here to
learn about the difference between a steady state
and a true equilibrium.

Cilia and Flagella

To understand the regulation of microtubule
assembly and function in any organism is a difficult
task. To study microtubules in cells as complex
vertebrate (e.g., human) cells is a nearly impossible
task, without a few "hints" as to how
to proceed. The basic mechanisms can be worked
out using a much less complex biological model such
as a flagellate. For example, the flagella of
the photosynthetic protist Chlamydomonas are
composed of microtubules, as are all flagella
and cilia.

Cilia and flagella have the same basic structure.
They are attached to structures known as basal
bodies, which in turn are anchored to the
cytoplasmic side of the plasma membrane. From
the basal bodies the microtubule "backbone" extends,
pushing the plasma membrane out with it.

To form cilia or flagella, microtubules arrange
themselves in a "9 + 2" array. Each of the two
central microtubules consists of a single microtubule
with 13 protofilaments arranged to form the wall
of a circular tube. Each of the outer nine consists
of a pair of microtubules that share a common
wall (see the cross sections of microtubules
in the figure). Look at the complete cross section
carefully. The hair-like appearance of flagella
and cilia in a light microscope is misleading.
The entire structure lies within the cytoplasm
of the cell.

The treatment given here to the structure of
microtubules ignores their true complexity. Functional
microtubules include associated proteins, anchoring
sites and organizing centers, sites for enzyme
activity, etc. In cilia and flagella, tubulin
forms a core structure to which other proteins
contribute structures called dynein arms, radial
spokes, and nexin links. The arms, spokes, and
links hold microtubules together and allow interaction
between microtubules that is superficially similar
to the sliding of actin and myosin filaments
in muscle contraction.

Ciliary and Flagellar Motion

One might appreciate the complexity of microtubular
organelles by looking at the motion of cilia
and flagella. Despite the similarities in structure,
the difference in nature of motility by flagella
versus cilia is profound, as one can see by comparing
representatives of the groups Ciliophora (the
ciliates) and Mastigophora (the flagellates).
Ciliates and flagellates behave differently,
live in different habitats and occupy different
niches, and likely represent two different evolutionary
lineages. The main difference in function is
in how they are organized.

Flagella are much longer than cilia and are
usually present singly or in pairs. A single
flagellum may propel the cell with a whip-like
motion. A pair of flagella may move in a synchronized
manner to pull the organism through the water,
in a way similar to the breast stroke of a human
swimmer.

Cilia tend to cover the surface area of a cell.
Both cilia and flagella bend as the microtubules
slide past one another. The arrangement of cilia
permits their coordinated movement in response
to signals from the cytoplasm. A small ciliate
may have hundreds of individual cilia, all beating
in a coordinated manner. How is all of the sliding
and bending coordinated? How does the organism "decide" in
what direction to move, or how to turn, rotate,
or feed? How does it convey the information to
hundreds of cilia to bend in a certain way? Questions
of that nature are fascinating to cell biologists.
They are very difficult to address, because each
system is so complex. Nevertheless, with a genome
about a hundred times smaller than that of a human,
a typical protist is much easier to study than
a human cell.

The motion of an individual cilium or flagellum
superficially resembles that of an oar, in that
it sweeps through the medium with a power stroke that
propels the cell. Each power stroke and return
stroke involves perhaps thousands of chemical reactions.
There may be dozens of strokes per second, and
one action may involve thousands of cilia. You
may notice that ciliates respond very quickly to
obstacles or changes in their environment. It is
fascinating to speculate on just how they receive
information, process it, and deliver the signals
to the cilia to produce precise movement. From
the perspective of our relatively slow world, it
is also difficult to comprehend how so much can
go on in such a short time. Think of the effect
on your perception of the universe if you were
to shrink by several orders of magnitude.

Copyright
and Intended Use
Visitors: to ensure that your message is not mistaken for
SPAM, please include the acronym "Bios211" in the subject line
of e-mail communications
Created by David R. Caprette (caprette@rice.edu), Rice University Dates